TRANSPORT AND METABOLISM OF THIAMINE IN RAT BRAIN CORTEX IN VITRO.

1. Aerobic incubation at 37 degrees of rat brain-cortex slices in Krebs-Ringer phosphate medium containing glucose and labelled thiamine results in accumulation in the tissue of labelled thiamine and labelled thiamine phosphates. The concentration of the labelled thiamine in the tissue cell water increases with increase of external labelled thiamine concentration in an approximately linear manner, the concentration ratio for labelled thiamine (tissue:medium) exceeding unity with low external thiamine concentrations (e.g. 0.2mum) and diminishing to about unity as the external thiamine concentration is increased to 1mum. The concentration of labelled phosphorylated thiamine in the tissue is at least double that of the labelled thiamine present and its amount increases with increase of external thiamine concentration. Labelled phosphorylated thiamine appears in the medium, its amount being about one-fifteenth of that in the tissue. Phosphorylation of thiamine in the tissue proceeds during incubation for 3hr. and, with an external labelled thiamine concentration of 0.2mum, about 48% conversion of thiamine takes place. 2. In the presence of ouabain (0.1mm), which does not inhibit thiamine phosphorylation in rat brain extract, there is a fall in the uptake of labelled thiamine by brain-cortex slices and the concentration ratio for the labelled thiamine (tissue:medium) falls to below unity. Anaerobiosis, lack of Na(+) or the presence of Amprol (0.01mm) leads to marked inhibition of thiamine phosphorylation, and the concentration ratio for labelled thiamine (tissue:medium) falls to about unity. The facts lead to the conclusion that thiamine is conveyed into the brain cell against a concentration gradient by an energy-assisted process mediated by a membrane carrier. Pyri-thiamine is a marked inhibitor of thiamine phosphorylation in brain extract. 3. Thiamine monophosphate and thiamine diphosphate inhibit thiamine phosphorylation in brain extract. They diminish ;total' thiamine (free and phosphorylated) uptake into brain-cortex slices and inhibit the transport of thiamine into the brain cell, possibly by competition for the carrier. 4. Phosphorylation of labelled thiamine in brain extract is brought about not only by adenosine triphosphate (in the presence of Mg(2+)) but apparently by adenosine diphosphate and uridine triphosphate.